PROJECT ABSTRACT Regulatory T (Treg) cells represent key cellular players in the immune response, tasked with maintaining balance in immunity. Endowed with potent suppressive function, Treg cells maintain immune homeostasis, prevent autoimmunity, and curtail robust immune responses to prevent collateral tissue damage. These cells are critically important, and their insufficiency or loss results in autoimmunity. However, they can also be over- activated, preventing effective immunity to chronic viral infections and cancer. We and others hypothesize that Treg cells differ from conventional T cells not only in their function, but also in how they fuel that function. The intersection of metabolism and immunity has been of immense recent interest; our laboratory studies how energetics impacts immune cell function with a focus on immunity in the tumor microenvironment. We hypothesize that, as regulatory T cells exist out-of-sync with their conventional counterparts, suppressing autoimmunity or curtailing productive immunity in the tissues, they exhibit metabolic plasticity, able to persist and thrive on alternative metabolic substrates in the environment. Our preliminary studies support this hypothesis, using cancer as a model. Tumor-infiltrating Treg cells are extraordinarily active and proliferative, and maintain their epigenetic and lineage stability despite substantial inflammatory insults and thus represent a model of an exemplary regulatory cell. We have found that Treg cells `stratify' into distinct metabolic subsets with different suppressive capacities. This New Innovator project seeks to understand how the microenvironment, in cancer and in other diseases, promotes Treg cell function through metabolic symbiosis. We present evidence of alternative metabolic pathways utilizing byproducts of tumor cell metabolism that may support Treg cell function in the tissues. We will explore, based on transcriptomic profiles of these cells, which metabolites present in the tumor microenvironment support Treg cell function and epigenetic stability. Further, we have developed a novel system by which certain metabolic pathways can be enforced in a Treg cell-specific manner. Taken together, these studies will shed light on how these crucial cells are exploited by cancer to evade the immune response but also open up potential avenues by which these cells may be therapeutically modulated through metabolic means to prevent autoimmunity or promote the anti-tumor response in cancer.